The long term goal of this proposal is to define the role of insulin like growth factor binding protein-2 (IGFBP-2) in skeletal acquisition and maintenance, and to determine the mechanisms whereby this molecule acts synergistically with IGF-I to regulate bone remodeling. The IGF regulatory system in bone includes a family of highly conserved IGF binding proteins (IGFBPs), proteases, receptors and the IGFs. IGFBP-2 is highly expressed in osteoblasts, and it stimulates mesenchymal stromal cell (MSC) proliferation. Global Igfbp2-/- mice have low bone turnover, impaired osteoblast and osteoclast differentiation, and accelerated marrow adipogenesis. To determine the mechanism of action of IGFBP-2 independent of IGF-I, we synthesized a short 13 amino acid peptide that does not bind IGFs but contains a unique heparin binding domain within IGFBP-2 (i.e. HBD1), and showed that HBD1 stimulated OB differentiation as did intact IGFBP-2. Daily administration of a pegylated form of the HBD1 peptide to Igfbp2 -/- mice for 3 weeks rescued their low trabecular bone mass, increased OB number, and enhanced hematopoietic progenitor cells. Osteoblasts from Igfbp2-/- mice have increased PTEN (phosphatase and tensin homolog) transcripts which can be suppressed by addition of exogenous IGFBP-2, IGF-I or HBD. Phosphorylation of AKT in MSCs from Igfbp2-/- mice was greatest when IGF-I + IGFBP-2 were added. Taken together, we hypothesize that IGFBP-2, which is induced by IGF-I, stimulates MSCs and OBs through HBD1. Furthermore we postulate that IGF-I and IGFBP-2 normally act in a complementary manner to promote bone remodeling by acting synergistically through distinct cell surface receptors. The two specific aims proposed are: 1) to determine IGFBP-2 actions on bone remodeling and its role in lineage allocation of MSCs, HSCs, and osteoclast precursors. 2) To delineate the molecular mechanism(s) of IGFBP-2 actions on bone cells, and determine if HBD1 is sufficient to mediate these effects through the pleiotrophin receptor. Using the HBD1 peptide we will study how IGFBP-2 affects the intracellular signaling pathways that promote cell proliferation. These interdisciplinary studies will lead to a clearer understanding of the function of IGFBP-2 in the adult skeleton. PUBLIC HEALTH RELEVANCE: This proposal has significant public health implications since it will help delineate the role of the IGF regulatory system in bone. Importantly, studies of the heparin binding domain will determine whether this small molecule can be used for the treatment of metabolic bone disorders.